Previous studies in cultured human fibroblasts have identified the events in the natural history of a typical lysosomal enzyme: synthesis of the polypeptide as a precursor of higher molecular weight, glycosylation in the endoplasmic reticulum, phosphorylation in the Golgi, receptor mediated translocation to lysosomes where limited proteolysis yields the mature enzyme or alternatively, secretion of the modified precursor and subsequent receptor-mediated endocytosis. This sequence was worked out by biosynthetic labeling of cultured fibroblasts followed by isolation of several enzymes by immunoprecipitation and gel electrophoresis. In the case of the lysosomal enzyme beta-hexosaminidase, there is another critical event - i.e., association of the alpha and beta subunits to give the catalytically active A isozyme. Using antibodies that distinguish between free and bound forms of the alpha chain we found that subunit association occurs relatively late in the course of biosynthesis, that it is essential for transfer of the alpha chains to lysosomes and that it requires protein synthesis, perhaps to provide a continuous supply of beta chains. Lack of association of the two subunits has been observed in cells from a number of patients with nonclassical (juvenile, adult and chronic) forms of Tay-Sachs disease, sometimes superimposed on a reduced rate of synthesis of the alpha chain. In all cases there was little or no detectable mature (lysosomal) form of the alpha chain. This differs from findings in classical (late infantile) Tay-Sachs disease, which may be caused by total lack of alpha chain synthesis (Ashkenazi variant) or by the synthesis of a defective alpha chain. Lack of synthesis of beta chains has been noted in two instances of Sandhoff disease. We are cloning cDNA for the alpha and beta chains of beta-hexosaminidase in order to investigate the precise nature of these mutations.